Putative HIV-1 reverse transcriptase inhibitors: design, synthesis, in vitro evaluation and in silico analysis.
Loading...
Date
2018
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
Abstract
One of the most significant treatments for HIV-1 infection has been the combination of drugs
targeting the HIV life cycle with the aim of preventing further destruction of the host immune
system. This study addresses the design, synthesis, in vitro evaluation, and in silico analysis of
putative HIV-1 reverse transcriptase (RT) inhibitors. The inhibitors comprise two structurally
diverse components which are intended to bind separately to the enzyme allosteric site and to
a location at, or close to, the polymerase active site. Therefore, the hydrophobic N-tritylated phalo-DL-phenylalanine derivatives (fluoro, chloro, bromo, iodo) have been coupled to 8-(6-
aminohexyl) amino-adenosine-3',5'-cyclic monophosphate through N-hydroxysuccinimidecarbodiimide chemistry.
Compounds were characterized by thin layer chromatography, UV spectroscopy, MALDI-TOF
mass spectrometry and proton NMR spectrometry. A reverse transcriptase colorimetric assay
kit, which features a sandwich ELISA protocol, based on biotin-avidin and digoxygenin-anti
DIG interactions, was used for quantitative determination of the inhibitory effect of synthesized
compounds on recombinant HIV-1 reverse transcriptase activity in vitro. Molecular docking
simulations of the chimeric inhibitors within the allosteric site of HIV-1 RT, were performed
using AutoDock Vina. The predicted binding associations were compared with laboratory
findings on HIV-1 RT inhibition. Two dimensional representations of protein-ligand
interactions were generated using LigPlot.
The non-halogenated N-trityl-L-phenylalanine-8-(6-aminohexyl)amino-adenosine-3',5'-cyclic
monophosphate derivative (4a) inhibited RT activity down to 57 % at 10-4 M, while the Ntrityl-para-fluoro-DL-phenylalanine-8-(6-aminohexyl)aminoadenosine-3′,5′-cyclic
monophosphate derivative (4b) was the strongest RT inhibitor reducing RT activity to 69 % at
10-7 M (IC50 = 29.2 μM). In the same assay, Nevirapine, a first-line anti-retroviral drug, showed
a decline in RT activity down to 43% at 10-5 M (IC50 = 3.03 μM).
Ranking of inhibitors according to estimated docking energies obtained from in silico docking
was in excellent agreement with potencies calculated from experimental studies. The docking
score of N-trityl-para-fluoro-DL-phenylalaline-8-(6-aminohexyl)amino-adenosine-3',5'-cyclic
monophosphate was -8.8 kcal/mol, while that of Nevirapine was -9.9 kcal/mol. The benzene
rings of the N-trityl-fluoro-DL-phenylalanine-8-(6-aminohexyl) amino-adenosine-3',5'-cyclic
monophosphate derivative formed hydrophobic interactions with hydrophobic, non-aromatic
amino acid residues Pro176 and Val179 in the allosteric site. Nevirapine, on the other hand
showed strong van der Waals interactions with Val106 ,Val179 and Tyr188 due to the aromatic
properties of the pyridine ring. Possible π-π stacking between phenyl rings of Nevirapine and
Tyr 181/Tyr188 aromatic side chains may also be present. Other HIV-1 RT large subunit
residues in the allosteric site common to the binding of Nevirapine and the active para-fluoro
derivative include Lys101, Tyr318, Leu 100, Trp229 and Phe227. Apparent binding to the
allosteric site suggests that compounds may be acting primarily as non-nucleoside reverse
transcriptase inhibitors (NNRTIs).
Description
Masters Degree. University of KwaZulu-Natal, Durban.